The broad and long-term objective of this project has been and continues to be the elucidation of the structure function relationships in selected salivary molecules. The biophysical methodologies employed to study these macromolecules will provide specific information as to the molecular nature of their biological behavior. Such data is essential if one wishes to understand the processes governing both the normal and diseased states in the oral cavity. As part of our continuing studies. three salivary molecules with demonstrated biological activities will be examined. The first of these is the proline-rich glycoprotein from human parotid saliva (PRG). The biological functions of PRG include masticatory lubrication, bacterial binding, pellicle formation and calcium coordination. Secondly we will continue studying a tyrosine-rich phosphoprotein called statherin. The primary biological function of statherin is the regulation of the calcium-phosphate equilibrium between saliva and the tooth. Finally, the structure of the acidic proline-rich proteins (aPRP's) will be investigated. The aPRP's like stathrin are known to be calcium binding proteins. The biological function common to all three of these salivary molecules is that of calcium coordination. The molecular mechanism(s) and conformational changes occurring in these macromolecules required to bind calcium is largely unknown. Detailed structural data on the free and metal-bound salivary molecules as well as their bioactive constituents will be obtained. Initially, optical spectroscopic techniques (e.g. fluorescence, ultraviolet & visible, circular dichroism) will be used to discern bulk secondary and tertiary structures. High resolution nuclear magnetic resonance spectroscopy will then be used to elucidate the spatial orientations of these salivary molecules. Comparative spectroscopic investigations using 40Ca, appropriate rare earth metals, 113 Cd and 43Ca will be conducted to completely evaluate the nature of the metal binding site(s) in these molecules. Lastly, the collective data will be refined using computer modeling techniques with the internuclear distance and torsion angle constraints acquired from the spectroscopic studies. The results obtained from these data will provide specific information regarding metal protein stoichiometry, the interaction of the salivary molecules with themselves(e.g. duplex or higher order self aggregation), the interaction of the salivary molecules with themselves (e.g. duplex or higher order self-aggregation), the conformation(s) of the metal-free and metal-bound molecules, and the refined structures of the metal binding sites. The data will ultimately be collated to ascertain if these salivary molecules have a common method of calcium coordination. Thus, the results derived from these studies will provide the first correlations of biological activity with conformation in salivary molecules at atomic resolution.